Genomic surveillance elucidates Ebola virus origin and transmission during the 2014 outbreak

Abstract

In its largest outbreak, Ebola virus disease is spreading through Guinea, Liberia, Sierra Leone, and Nigeria. We sequenced 99 Ebola virus genomes from 78 patients in Sierra Leone to ~2000× coverage. We observed a rapid accumulation of interhost and intrahost genetic variation, allowing us to characterize patterns of viral transmission over the initial weeks of the epidemic. This West African variant likely diverged from central African lineages around 2004, crossed from Guinea to Sierra Leone in May 2014, and has exhibited sustained human-to-human transmission subsequently, with no evidence of additional zoonotic sources. Because many of the mutations alter protein sequences and other biologically meaningful targets, they should be monitored for impact on diagnostics, vaccines, and therapies critical to outbreak response.

Fig. 1. Ebola outbreaks, historical and current

(A) Historical EVD outbreaks, colored by decade. Circle area represents total number of cases (RC = Republic of the Congo; DRC = Democratic Republic of Congo). (B) 2014 outbreak growth (confirmed, probable, and suspected cases). (C) Spread of EVD in Sierra Leone by district. The gradient denotes number of cases; the arrow depicts likely direction. (D) EBOV samples from 78 patients were sequenced in two batches, totaling 99 viral genomes [replication = technical replicates (6)]. Mean coverage and median depth of coverage with range are shown. (E) Combined coverage (normalized to the sample average) across sequenced EBOV genomes.

Fig. 2. Relationship between outbreaks

(A) Unrooted phylogenetic tree of EBOV samples; each major clade corresponds to a distinct outbreak (scale bar = nucleotide substitutions per site). (B) Root-to-tip distance correlates better with sample date when rooting on the 1976 branch (R² = 0.92, top) than on the 2014 branch (R² = 0.67, bottom). (C) Temporally rooted tree from (A).

Fig. 3. Molecular dating of the 2014 outbreak

(A) BEAST dating of the separation of the 2014 lineage from central African lineages [SL, Sierra Leone; GN, Guinea; DRC, Democratic Republic of Congo; time of most recent common ancestor (tMRCA), September 2004; 95% highest posterior density (HPD), October 2002 to May 2006]. (B) BEAST dating of the tMRCA of the 2014 West African outbreak (23 February; 95% HPD, 27 January to 14 March) and the tMRCA of the Sierra Leone lineages (23 April; 95% HPD, 2 April to 13 May). Probability distributions for both 2014 divergence events are overlaid below. Posterior support for major nodes is shown.

Fig. 4. Viral dynamics during the 2014 outbreak

(A) Mutations, one patient sample per row; beige blocks indicate identity with the Kissidougou Guinean sequence (GenBank accession KJ660346).The top row shows the type of mutation (green, synonymous; pink, nonsynonymous; gray, inter-genic), with genomic locations indicated above. Cluster assignments are shown at the left. (B) Number of EVD-confirmed patients per day, colored by cluster. Arrow indicates the first appearance of the derived allele at position 10,218, distinguishing clusters 2 and 3. (C) Intrahost frequency of SNP 10,218 in all 78 patients (absent in 28 patients, polymorphic in 12, fixed in 38). (D and E) Twelve patients carrying iSNV 10,218 cluster geographically and temporally (HCW-A = unse-quenced health care worker; Driver drove HCW-A from Kissi Teng to Jawie, then continued alone to Mambolo; HCW-B treated HCW-A). KGH = location of Kenema Government Hospital. (F) Substitution rates within the 2014 outbreak and between all EVD outbreaks. (G) Proportion of nonsynonymous changes observed on different time scales (green, synonymous; pink, nonsynonymous). (H) Acquisition of genetic variation over time. Fifty mutational events (short dashes) and 29 new viral lineages (long dashes) were observed (intrahost variants not included).